A method is described for measuring the potential energy of nanoparticles in an optical trap by trapping an ensemble of particles with a focused laser beam. The force balance between repulsive osmotic and confining gradient-force pressures determines the single-particle trapping potential independent of interactions between the particles. The ensemble nature of the measurement permits evaluation of single-particle trapping energies much smaller than kBT. Energies obtained by this method are compared to those of single-particle methods as well as to theoretical calculations based on classical electromagnetic optics.